JP6751475B2 - Pharmaceutical composition for sublingual administration of edaravone (+)-2-borneol - Google Patents

Description

The present invention belongs to the technical field of medicine, and relates to a pharmaceutical composition for sublingual administration of edaravone.(+)-2-borneol, and a method for producing the same.

Edaravone (chemical name: 3-methyl-1-phenyl-2-pyrazolin-5-one) is a brain protective agent (YakugakuZasshi.2004,124(3):99-111) which is already on the market. Studies show that edaravone has antioxidant activity, significantly improves the neurological deficit symptoms of cerebral ischemia-reperfusion, reduces infarct size, reduces the extent of brain damage and reduces cerebral edema. , Can suppress lipid peroxidation in damaged brain tissue.

(+)-2-Bornole is the main component of natural borneol, which is a commonly used Chinese medicine, and borneol has the effects of "resuscitation opening", "aromatic movement", and "going to the medicine". It is often used as a "menopausal drug" so as to enhance the therapeutic effect of the drug. In "Bencao Yanyi, Augmented Materia Medica", borneol has a weak momentum when used alone, but instead has a pharmacodynamic effect when used as an adjuvant. Animal studies and in vitro studies have shown that borneol has the effect of promoting drug penetration into the blood-brain barrier.

Since cerebrovascular disease, particularly ischemic cerebrovascular disease, is an acute disease and it is necessary to rapidly relieve symptoms, injection is the first choice for emergency medical care. The invention patent (CN101848711A), whose name is “Pharmaceutical composition and its use in the preparation of a medicament for treating cerebrovascular disease”, describes that a composition of edaravone and (+)-2-borneol in a specific proportion of an injectable solution is brain. It discloses the application to the preparation of a medicament for the treatment of vascular diseases, in particular ischemic cerebrovascular diseases, and discloses a better therapeutic effect of the composition compared to edaravone injection.
However, intramuscular or intravenous injection not only causes pain and irritation at the injection site and needs to be manipulated by medical staff, but it also requires injection articles and the like, which is a practical medical constraint. It is not suitable for patients who have been treated outside the hospital.

The preparation for sublingual administration is directly absorbed from the sublingual mucosa, which has a large surface area and high permeability, and a large amount of capillaries from the submucosa gather in the internal jugular vein and pass through the superior vena cava. After entering the blood circulation and after administration, the drug is rapidly absorbed, resulting in rapid effect, accurate quantification, convenient to use, avoiding the first-pass effect of orally administered drug. Compared with injection, sublingual tablets can greatly improve the convenience of drug administration and the compliance of clinical patients.
However, a pharmaceutical composition for sublingual administration containing edaravone and (+)-2-borneol is not easily prepared. The present inventor has found that general excipients cannot produce a sublingual tablet that has passed, or that the performance (stability, release rate, etc.) of the produced sublingual tablet cannot be satisfied. It was
Therefore, even in the prior art, a pharmaceutical composition for sublingual administration containing edaravone and (+)-2-borneol, which has satisfactory stability and release rate, is urgently desired.

CN101848711A

YakugakuZasshi.2004,124 (3): 99-111

An object of the present invention is to provide a pharmaceutical composition for sublingual administration containing edaravone or a salt thereof and (+)-2-borneol.
The volatility of (+)-2-borneol makes it difficult to stabilize the (+)-2-borneol content in solid formulations, and the drug in sublingual tablets can be released rapidly and consistently. The therapeutic effect can be exerted only when the blood concentration of the drug is reached. The present inventor has unexpectedly discovered that the above problem can be effectively solved by using an excipient containing a combination of mannitol and copolyvidone. Based on the above findings, the present invention, in the pharmaceutical composition for sublingual administration containing edaravone or a salt thereof, and (+)-2-borneol, the content of (+)-2-borneol is stable, Further, the present invention provides a pharmaceutical composition for sublingual administration, in which the pharmaceutically active ingredient is rapidly released and absorbed under the tongue.

In order to achieve the above object, the present invention employs the following technical solutions.

It contains edaravone or a salt thereof and (+)-2-borneol as an active ingredient, and an excipient, and a pharmaceutically acceptable additive, wherein the excipient is mannitol, lactose, dextran, cysteine, glycine. Edaravone and (+)-2-borneol, characterized in that it is one or more selected from copolyvidone and β-cyclodextrin, and preferably the excipient contains a combination of mannitol and copolyvidone. A sublingual tablet containing the composition of.

The preferred pharmaceutical composition for sublingual administration of the present invention contains mannitol and copolyvidone as excipients, and their mass ratio is 1:5 to 5:1, preferably 1:1 to 5:1.

In some preferred embodiments of the invention, the excipient comprises a combination of mannitol, copolyvidone, and microcrystalline cellulose. Preferably, the mass ratio of mannitol, copolyvidone, and microcrystalline cellulose is (5-10):(1-5):(10-20).

In some other preferred embodiments of the invention, the excipient comprises a combination of mannitol, copolyvidone, and lactose. Preferably, the mass ratio of mannitol, copolyvidone and lactose is (1-10):(1-5):(10-30).

In the pharmaceutical composition for sublingual administration, the mass ratio of edaravone to (+)-2-borneol as a free base is greater than 4 or less than 1, preferably edaravone and (+)-as a free base. The mass ratio with 2-borneol is more than 4 and less than 10, or more than 0.1 and less than 1, and more preferably, the mass ratio of edaravone and (+)-2-borneol as a free base is 5.

In the pharmaceutical composition for sublingual administration, the weight ratio of (+)-2-borneol to the excipient is 0.1 to 1, preferably 0.3 to 1, and more preferably 0.3 to 0.5.

The method for preparing the pharmaceutical composition for sublingual administration is as follows: (+)-2-borneol is dissolved in an ethanol solution, an excipient is dissolved in an aqueous solution, both are combined and stirred, left to stand, and freeze-dried. , Sieving, adding edaravone and other additives, mixing uniformly and tableting.

In the pharmaceutical composition for sublingual administration, the drug blood concentration of edaravone reaches 10 to 8000 ng/mL within about 0.1 to 10 hours after the administration of the preparation of each standard unit, and (+)-2-borneol. The drug blood concentration of edaravone reaches 1 to 200 ng/mL, preferably within about 0.1 to 6 hours after administration of the formulation per standard unit, and the drug blood concentration of edaravone reaches 50 to 5000 ng/mL. , (+)-2-Bornole drug concentration in blood reaches 2-150 ng/mL.

FIG. 1 is an elution curve of Examples 1-5. FIG. 2 is an elution curve of Examples 6-11. FIG. 3 is an elution curve of Examples 8 and 12-14.

The present invention discloses a formulation for sublingual administration of edaravone and (+)-2-borneol, and a person skilled in the art, while referring to the content of the present invention, can combine the principles of pharmaceutics and appropriately process parameters. Alternatively, it can be achieved by improving the compounding ratio in the formulation. It should be pointed out that all similar substitutions and modifications will be apparent to those skilled in the art, and all of them fall within the scope of the present invention. The application of the invention is described by means of preferred embodiments, and it will be apparent to those skilled in the art that modifications or appropriate changes to the methods and applications described herein may be made without departing from the scope, spirit and scope of the disclosure. It is obvious that the technology of the present invention can be realized and applied by combining the above and the combination.

The edaravone described in the examples is 3-methyl-1-phenyl-2-pyrazolin-5-one.
Hereinafter, the present invention will be further described with reference to examples, but the present invention is not limited to the examples.

調製方法は、（+）-2-ボルネオール、エダラボン、ラクトース、ヒプロメロース、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを均一に混合し、打錠した。 Example 1

The preparation method was as follows: (+)-2-borneol, edaravone, lactose, hypromellose, cross-linked sodium carboxymethyl cellulose, magnesium stearate were uniformly mixed and tableted.

調製方法は、（+）-2-ボルネオール、エダラボン、ラクトース、ヒプロメロース、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを均一に混合し、打錠した。 Example 2

The preparation method was as follows: (+)-2-borneol, edaravone, lactose, hypromellose, cross-linked sodium carboxymethyl cellulose, magnesium stearate were uniformly mixed and tableted.

調製方法は、（+）-2-ボルネオール、エダラボン、ラクトース、ヒプロメロース、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを均一に混合し、打錠した。 Example 3

The preparation method was as follows: (+)-2-borneol, edaravone, lactose, hypromellose, cross-linked sodium carboxymethyl cellulose, magnesium stearate were uniformly mixed and tableted.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、β-シクロデキストリンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、ラクトース、ヒプロメロース、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 4

The preparation method is as follows: (+)-2-borneol is dissolved in an ethanol solution, β-cyclodextrin is dissolved in an aqueous solution, both of them are stirred together, left to stand, freeze-dried, sieved, edaravone, lactose, Hypromellose, cross-linked sodium carboxymethyl cellulose and magnesium stearate were added and mixed uniformly and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、β-シクロデキストリンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、ラクトース、ヒプロメロース、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 5

The preparation method is as follows: (+)-2-borneol is dissolved in an ethanol solution, β-cyclodextrin is dissolved in an aqueous solution, both of them are stirred together, left to stand, freeze-dried, sieved, edaravone, lactose, Hypromellose, cross-linked sodium carboxymethyl cellulose and magnesium stearate were added and mixed uniformly and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、（+）-2-ボルネオールの重量に対して5倍量のマンニトール、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、残りのマンニトール、コポリビドン、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 6

The preparation method is to dissolve (+)-2-borneol in an ethanol solution, dissolve 5 times the amount of mannitol and a small amount of copolyvidone in the aqueous solution, and stir them together. Then, the mixture was allowed to stand, freeze-dried, sieved, edaravone, the remaining mannitol, copolyvidone, cross-linked sodium carboxymethyl cellulose, and magnesium stearate were added, and the mixture was uniformly mixed and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、（+）-2-ボルネオールの重量に対して3倍量のマンニトール、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、残りのマンニトール、コポリビドン、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 7

The preparation method is to dissolve (+)-2-borneol in an ethanol solution, dissolve 3 times the amount of mannitol and a small amount of copolyvidone in the aqueous solution, and stir them together. Then, the mixture was allowed to stand, freeze-dried, sieved, edaravone, the remaining mannitol, copolyvidone, cross-linked sodium carboxymethyl cellulose, and magnesium stearate were added, and the mixture was uniformly mixed and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、マンニトール、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、ラクトース、コポリビドン、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 8

The preparation method is as follows: (+)-2-borneol is dissolved in ethanol solution, mannitol and a small amount of copolyvidone are dissolved in aqueous solution, both are combined and stirred, left to stand, freeze-dried, sieved, edaravone, lactose. , Copolyvidone, cross-linked sodium carboxymethyl cellulose, and magnesium stearate were added and uniformly mixed and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、マンニトール、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、ラクトース、コポリビドン、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 9

The preparation method is as follows: (+)-2-borneol is dissolved in ethanol solution, mannitol and a small amount of copolyvidone are dissolved in aqueous solution, both are combined and stirred, left to stand, freeze-dried, sieved, edaravone, lactose. , Copolyvidone, cross-linked sodium carboxymethyl cellulose, and magnesium stearate were added and uniformly mixed and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、（+）-2-ボルネオール重量に対して5倍量のラクトース、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、残りのラクトース、コポリビドン、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 10

The preparation method is to dissolve (+)-2-borneol in an ethanol solution, and to dissolve 5 times the amount of lactose and a small amount of copolyvidone in the aqueous solution relative to the weight of (+)-2-borneol, and stir them together. The mixture was allowed to stand, freeze-dried, sieved, edaravone, the remaining lactose, copolyvidone, cross-linked sodium carboxymethyl cellulose, and magnesium stearate were added and uniformly mixed to give tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、（+）-2-ボルネオール重量に対して1.25倍量のラクトース、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、残りのラクトース、コポリビドン、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 11

The preparation method is as follows: (+)-2-borneol is dissolved in an ethanol solution, 1.25 times the amount of lactose and a small amount of copolyvidone are dissolved in the aqueous solution, and both are combined and stirred. The mixture was allowed to stand, freeze-dried, sieved, edaravone, the remaining lactose, copolyvidone, cross-linked sodium carboxymethyl cellulose, and magnesium stearate were added and uniformly mixed to give tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、マンニトール、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、微結晶セルロース、コポリビドン、架橋カルボキシメチルセルロースナトリウム、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 12

The preparation method is as follows: (+)-2-borneol is dissolved in ethanol solution, mannitol and a small amount of copolyvidone are dissolved in aqueous solution, and both are stirred together, left to stand, freeze-dried, sifted, edaravone, fine Crystalline cellulose, copolyvidone, cross-linked sodium carboxymethyl cellulose and magnesium stearate were added and uniformly mixed and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、マンニトール、コポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、微結晶セルロース、架橋カルボキシメチルセルロースナトリウム、シリカ、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 13

The preparation method is as follows: (+)-2-borneol is dissolved in ethanol solution, mannitol and copolyvidone are dissolved in aqueous solution, both are combined and stirred, left to stand, freeze-dried, sieved, edaravone, microcrystalline cellulose , Cross-linked sodium carboxymethyl cellulose, silica, and magnesium stearate were added and uniformly mixed, and compressed into tablets.

調製方法は、（+）-2-ボルネオールをエタノール溶液に溶解させ、マンニトール、少量のコポリビドンを水溶液に溶解させ、両方を合わせて撹拌し、静置し、凍結乾燥し、篩にかけ、エダラボン、微結晶セルロース、コポリビドン、架橋カルボキシメチルセルロースナトリウム、シリカ、ステアリン酸マグネシウムを加えて均一に混合し、打錠した。 Example 14

The preparation method is as follows: (+)-2-borneol is dissolved in ethanol solution, mannitol and a small amount of copolyvidone are dissolved in aqueous solution, and both are stirred together, left to stand, freeze-dried, sifted, edaravone, fine Crystalline cellulose, copolyvidone, cross-linked sodium carboxymethyl cellulose, silica, and magnesium stearate were added and uniformly mixed and compressed into tablets.

上記のデータにより、実施例の安定性試験の結果から、各実施例4〜14は、高温で30日置かれ、安定性がいずれも良く、そのうち、実施例8、12、13、14は高温で90日置かれ、安定性が比較的に良いことが示された。 Example 15
The stability test results were obtained by taking an appropriate amount of the samples of Examples 1 to 14, imitating the packaging on the market, and sampling at 10 days, 30 days, and 90 days under the conditions of temperatures of 40° C. and 60° C., and The table below shows the results of measurement of properties, content, and related substances.

According to the above data, from the results of the stability test of Examples, each of Examples 4 to 14 was placed at a high temperature for 30 days and had good stability, among which Examples 8, 12, 13, and 14 were at a high temperature. After 90 days, it was shown to be relatively stable.

Example 16
The elution rate measurement test method was to take out a sample and use 900 ml of water as an elution medium according to the elution rate/release rate measurement method described in “People's Republic of China Pharmacopoeia” 2015 Edition (Part 4, 0931, Second Method). Here, in Examples 3 and 4, 250 ml of water was used as the elution medium), operated at a rotation speed of 50 rpm, sampled at different time points, filtered through a 0.8 μm filtration membrane, and the filtrate was used as a sample solution, and a control An appropriate amount of edaravone to be used as a product was taken out, 20 mmol/L ammonium acetate/acetonitrile (80:20) was added and dissolved, and diluted to about 0.02 mg/ml to prepare. The UV absorbance of the sample solution was measured under the condition of 254 nm, and the elution rate of the sample was calculated. The results are shown in FIGS.

Example 17
Study on the distribution of edaravone sublingual tablets in plasma and brain tissue of SD rats
1. Material and method
1.1 Laboratory animals
Sprague-Dawley (SD) rat, SPF grade, male, weight 180-200g.
Origin: Beijing Weitong Lihua Experimental Animal Technology Co., Ltd.
Pass certificate number: 11400700138404.
License number: SCXK (Kyoto) 2012-0001.
Food and water supply: Before the experiment, the animals were fasted for 12 hours, but food was provided from 4 hours after the administration, and the experiment was not performed without water. Abnormal reactions in animals were observed and recorded during the administration and sampling process.

1.2 Reagent products Complex edaravone injection: Standard 12.5 mg/5 mL (edaravone and (+)-2-borneol are 10 mg/5 mL and 2.5 mg/5 mL, respectively).
Sublingual tablets were prepared at the ratios in the formulation of Example 8, and each tablet contained 5 mg of edaravone and 1 mg of borneol.

1.3 Method Group 1: Administration of compound edaravone injection (N=4) by intravenous injection Dose: edaravone 16 mg/kg, (+)-2-borneol 4 mg/kg, administration volume 8 mL/kg. Sampling time points for plasma and brain tissue samples: 2 min, 15 min, 30 min, 1 h, 2.5 h, 5 h. At each time point, whole blood and brain tissue were collected at the same time.
Group 2: Sublingual administration of one sublingual tablet (N=6).
As SD rats are in a lightly anesthetized state, chloral hydrate is intraperitoneally injected, the rat's oral cavity is moistened with 50 μL of water, and a sublingual tablet is placed under the rat's tongue, 1 tablet/mouse rat, The mouth of the rat was fixed for 30 minutes so that the tablets would not fall or slip into the gastrointestinal tract. The time for putting the tablets under the tongue was set to 0 min, and thereafter, whole blood and brain tissue were collected at 5 min, 15 min, 30 min, 1 h, 2.5 h, and 5 h, respectively.

SDラットの静脉注射による複合エダラボン注射液及び舌下錠の投与後の脳ホモジネート中のエダラボンの各薬物動態パラメータの平均値。

SDラットの静脉注射による複合エダラボン注射液及び舌下錠の投与後の血漿中のボルネオールの各薬物動態パラメータの平均値。

SDラットの静脉注射による複合エダラボン注射液、及び舌下による舌下錠の投与後の脳ホモジネート中のボルネオールの各薬物動態パラメータの平均値。

2. Experimental results
Mean value of each pharmacokinetic parameter of edaravone in plasma after administration of compound edaravone injection and sublingual tablet by intravenous injection in SD rat.

Mean values of each pharmacokinetic parameter of edaravone in the brain homogenate after administration of the compound edaravone injection solution and sublingual tablet by intravenous injection in SD rats.

Mean values of the respective pharmacokinetic parameters of borneol in plasma after administration of the compound edaravone injection by substitial injection and sublingual tablets in SD rats.

Mean value of each pharmacokinetic parameter of borneol in brain homogenate after administration of combined edaravone injection by intravenous injection and sublingual tablet in SD rat.

The results of studies on the distribution of the sublingual tablets in plasma and brain tissue of SD rats show that the sublingual formulation of edaravone and (+)-2-borneol has a bioavailability of about (62.6%, edaravone, 51.6%). %, (+)-2-borneol), and the bioavailability in the brain is about (28.5%, edaravone 28.5%, (+)-2-borneol), and sublingually administered edaravone. The bioavailability of and borneol was high, and it was shown that the sublingual administration condition was satisfied.
Sublingual tablets with edaravone and (+)-2-borneol have advantages such as good pharmacokinetic properties, high bioavailability, high blood-brain barrier permeability, convenient use of sublingual tablets.

Example 18
Protective effect of sublingual tablet administration on focal cerebral ischemia reperfusion injury
1 Materials and methods
1.1 Laboratory animals
Sprague-Dawley (SD) Rat, Male, Clean Grade, Weight 260-280g
1.2 Reagent product A sublingual tablet was prepared at the ratio in the formulation of Example 8, and edaravone 0.67 mg and borneol 0.13 mg (3 mg/kg dose group), edaravone 2.01 mg and borneol 0.39 mg (9 mg/kg dose group) , Edaravone 6mg and borneol 1.2mg (27mg/kg dose group) respectively, there are three standards in total.
The compound edaravone injection has a standard of 5mL: 12.5mg and was manufactured by Nanjing Singh Dongyuan Pharmaceutical Co., Ltd.

1.3 Experimental method
1.3.1 Preparation of focal cerebral ischemia-reperfusion model A cerebral ischemia-reperfusion model of middle cerebral artery occlusion (MCAO) was prepared by the internal carotid embolization method. The animals were anesthetized with 7% trichloroacetaldehyde monohydrate (6 ml/kg) and then fixed to the operating table in the prone position, the skin was disinfected, an incision was made in the middle of the neck, the right common carotid artery, Separate the external carotid artery and internal carotid artery, lightly dissect the vagus nerve, ligate and cut the external carotid artery, move forward along the internal carotid artery, and ligate the pterygopalatine artery (PPA) did. The proximal end of the common carotid artery is occluded, an incision is cut out at the distal end of the ligature of the external carotid artery, a nylon thread with an outer diameter of 0.285 mm is inserted, and the internal carotid artery is passed through the bifurcation of the common carotid artery. And gradually inject until there is a slight resistance (about 20 mm from the bifurcation), cut off all blood supply of the middle cerebral artery, and slowly slow down the nylon thread after 2.0 hours ischemia in the right brain. The blood supply was restored, reperfusion was performed, the skin was sutured and disinfected.

1.3.2 Grouping and administration of animals Experimental animals consisted of three sublingual tablets of the composition (3 mg/kg, 9 mg/kg, 27 mg/kg), a positive control drug complex edaravone group (3 mg/kg). ) And a model group, a total of 5 groups. After creating the cerebral ischemia model, the animals were divided into each group in a single blind manner with the same probability. The animals in the composition group were sublingually administered with a sublingual tablet of an appropriate standard at the time of re-irrigation, 1 tablet per rat, and the tablet dropped or was put into the gastrointestinal tract until the tablet was completely absorbed. The mouth of the rat was fixed so as not to slip off, the animals of the positive control drug group were administered once from the tail vein immediately after reperfusion, and the animals of the model group were injected with an equal volume of physiological saline. After assessing neurological deficit symptoms 24 hours after cerebral ischemia, the animals were sacrificed, the brain was removed, stained and photographed to determine the cerebral infarct area.

1.3.3 Scoring of neurological deficit symptoms and measurement of cerebral infarction area Neurological deficit symptoms were evaluated by a modified Bederson 5 point scoring system. A single-blind method was used to evaluate neurological deficit symptoms after cerebral ischemia in rats. That is, the experiment designer marks the animals according to the grouping, and the experimenter who scores the neurological deficit symptoms did not know the status of the grouping of the animals. After the evaluation was completed, the grader submitted the various marked scoring results to the designer. The designer disclosed the experiment and scored each animal in each group.

To measure the degree of cerebral infarction, after sacrificing the animal, decapitating the brain, removing the olfactory brain, cerebellum and lower brainstem, rinsing blood stains on the brain surface with physiological saline, and leaving it on the surface Absorbed water, placed at -20°C for 20 min, taken out, immediately cut vertically downward from the plane intersecting the line of sight, coronal section, and cutting out sections every 2 mm posteriorly. After incubating in TTC staining solution (37°C, 90 min), normal brain tissue was stained crimson, ischemic brain tissue appeared white, and after rinsing with physiological saline, the brain sections were rapidly washed from front to back. They lined up, absorbed the water remaining on the surface and took a picture.

The photographs were processed by image analysis software, and the volume corresponding to the left brain and the volume of infarcts were calculated by the following formula to determine the percentage of infarcts.
Infarct size calculation method:
V＝t (A1+A2+A3+………+An)
t is the thickness of the section and A is the infarct area.
% I = 100% x (VC-VL)/VC
%I is the percentage of infarct volume, VC is the brain volume on the control side (left hemisphere) and VL is the volume of the non-infarct region on the infarct side (right hemisphere).

2 experimental results
2.1 Effects on neurological deficit symptoms Table 1 shows the degree of neurological deficit symptoms in each group of animals. Compared with the model group, the three sublingual doses of the composition (3, 9, 27 mg/kg) and the positive control compound edaravone (3 mg/kg) all showed significant neurological deficit symptoms. Can be improved.
Table 3. Effects of sublingual combined drug of edaravone and (+)-2-borneol on neurological deficit symptoms

2.2 Effect on cerebral infarct area The effect on cerebral infarct area is shown in Table 2. All three sublingual doses of the composition (3, 9, 27 mg/kg) and the positive control compound edaravone reduce cerebral infarct area after animal ischemia-reperfusion compared to the model group be able to.
Table 2 Effect of edaravone and (+)-2-borneol combined drug for sublingual administration on cerebral infarct area

Example 19
Studies on the distribution of edaravone sublingual tablets in SD rat plasma and brain tissue:
3. Material and method
1.4 Experimental animals
Sprague-Dawley (SD) rat, SPF grade, male, weight 180-200g.
Origin: Beijing Weitong Lihua Experimental Animal Technology Co., Ltd.
Pass certificate number: 11400700138404.
License number: SCXK (Kyoto) 2012-0001.
Food and water supply: Before the experiment, the animals were fasted for 12 hours, but food was provided from 4 hours after the administration, and the experiment was not performed without water. Abnormal reactions in animals were observed and recorded during the administration and sampling process.
1.5 Reagent products Complex edaravone injection: Standard 12.5 mg/5 mL (edaravone and (+)-2-borneol are 10 mg/5 mL and 2.5 mg/5 mL, respectively).
Sublingual tablets were prepared according to the ratio in the formulation of Example 13, and each tablet contained 5 mg of edaravone and 1 mg of borneol.

1.6 Method Group 1: Administration of complex edaravone injection (N=4) by intravenous injection Dose: edaravone 16 mg/kg, (+)-2-borneol 4 mg/kg, administration volume: 8 mL/kg. Sampling time points for plasma and brain tissue samples: 2 min, 15 min, 30 min, 1 h, 2.5 h, 5 h. At each time point, whole blood and brain tissue were collected at the same time.
Group 2: Sublingual administration of one sublingual tablet (N=6).
As SD rats are in a lightly anesthetized state, chloral hydrate is intraperitoneally injected, the rat's oral cavity is moistened with 50 μL of water, and a sublingual tablet is placed under the rat's tongue, 1 tablet/mouse rat, The mouth of the rat was fixed for 30 minutes so that the tablets would not fall or slip into the gastrointestinal tract. The time for putting the tablets under the tongue was set to 0 min, and thereafter, whole blood and brain tissue were collected at 5 min, 15 min, 30 min, 1 h, 2.5 h, and 5 h, respectively.

SDラットの静脉による複合エダラボン注射液及び舌下錠の投与後の脳ホモジネート中のエダラボンの各薬物動態パラメータの平均値。

SDラットの静脉による複合エダラボン注射液及び舌下錠の投与後の血漿中のボルネオールの各薬物動態パラメータの平均値。

SDラットの静脉による複合エダラボン注射液、及び舌下による舌下錠の投与後の脳ホモジネート中のボルネオールの各薬物動態パラメータの平均値。

4. Experimental results
Mean value of each pharmacokinetic parameter of edaravone in plasma after administration of compound edaravone injection and sublingual tablet by intravenous administration of SD rats.

Mean value of each pharmacokinetic parameter of edaravone in brain homogenate after administration of combined edaravone injection and sublingual tablet by intravenous administration of SD rats.

Mean values of each pharmacokinetic parameter of borneol in plasma after administration of a complex edaravone injection and sublingual tablets by intravenous administration in SD rats.

Mean values of each pharmacokinetic parameter of borneol in brain homogenate after administration of combined edaravone injection by SD rat infusion and sublingual tablet by sublingual administration.

The results of studies on the distribution of the sublingual tablet in plasma and brain tissue of SD rats show that the sublingual formulation of edaravone and (+)-2-borneol has a bioavailability of about (79.9%, edaravone, 60.1%). %, (+)-2-borneol), and the bioavailability in the brain is about (30.1%, edaravone, 29.6%, (+)-2-borneol), and sublingually administered edaravone. The bioavailability with borneol was high, and it was shown that the sublingual administration condition was satisfied.
Sublingual tablets with edaravone and (+)-2-borneol have advantages such as good pharmacokinetic properties, high bioavailability, high blood-brain barrier permeability, convenient use of sublingual tablets.

Example 20
Protective effect of sublingual tablet administration on focal cerebral ischemia reperfusion injury
1 Materials and methods
1.1 Laboratory animals
Sprague-Dawley (SD) Rat, Male, Clean Grade, Weight 260-280g
1.2 Reagent products Sublingual tablets were prepared according to the ratio in the formulation of Example 13, edaravone 0.67 mg and borneol 0.13 mg (3 mg/kg dose group), edaravone 2.01 mg and borneol 0.39 mg (9 mg/kg dose group). , Edaravone 6mg and borneol 1.2mg (27mg/kg dose group) respectively, there are three standards in total.
The compound edaravone injection has a standard of 5mL: 12.5mg and was manufactured by Nanjing Singh Dongyuan Pharmaceutical Co., Ltd.

1.3 Experimental method
1.3.1 Preparation of focal cerebral ischemia-reperfusion model A cerebral ischemia-reperfusion model of middle cerebral artery occlusion (MCAO) was prepared by the cervical internal carotid artery embolization method. The animals were anesthetized with 7% trichloroacetaldehyde monohydrate (6 ml/kg) and then fixed to the operating table in the prone position, the skin was disinfected, an incision was made in the middle of the neck, the right common carotid artery, Separate the external carotid artery and internal carotid artery, lightly dissect the vagus nerve, ligate and cut the external carotid artery, move forward along the internal carotid artery, and ligate the pterygopalatine artery (PPA). did. The proximal end of the common carotid artery is occluded, an incision is cut out at the distal end of the ligature of the external carotid artery, a nylon thread with an outer diameter of 0.285 mm is inserted, and the internal carotid artery is passed through the bifurcation of the common carotid artery. Then, gradually inject until there is a slight resistance (about 20 mm from the bifurcation), cut off all blood supply of the middle cerebral artery, and after 2.0 hours of ischemia in the right brain, nylon thread was applied. Slowly withdrawn, blood supply restored, reperfusion was performed, skin was sutured and disinfected.

1.3.2 Grouping and administration of animals Experimental animals consisted of three sublingual tablets of the composition (3 mg/kg, 9 mg/kg, 27 mg/kg), a positive control drug complex edaravone group (3 mg/kg). ) And a model group, a total of 5 groups. After creating the cerebral ischemia model, the animals were divided into each group in a single blind manner with the same probability. The animals in the composition group were sublingually administered with a sublingual tablet of an appropriate standard at the time of re-irrigation, 1 tablet per rat, and the tablet dropped or was put into the gastrointestinal tract until the tablet was completely absorbed. The mouth of the rat was fixed so as not to slip off, the animals of the positive control drug group were administered once from the tail vein immediately after reperfusion, and the animals of the model group were injected with an equal volume of physiological saline. After assessing neurological deficit symptoms 24 hours after cerebral ischemia, the animals were sacrificed, the brain was removed, stained and photographed to determine the cerebral infarct area.

1.3.3 Scoring of neurological deficit symptoms and measurement of cerebral infarction area Neurological deficit symptoms were evaluated by a modified Bederson 5 point scoring system. A single-blind method was used to evaluate neurological deficit symptoms after cerebral ischemia in rats. That is, the experiment designer marks the animals according to the grouping, and the experimenter who scores the neurological deficit symptoms did not know the status of the grouping of the animals. After the evaluation was completed, the grader submitted the various marked scoring results to the designer. The designer disclosed the experiment and scored each animal in each group.

To measure the degree of cerebral infarction, after sacrificing the animal, decapitating the brain, removing the olfactory brain, cerebellum and lower brainstem, rinsing blood stains on the brain surface with physiological saline, and leaving it on the surface Absorbed water, placed at -20°C for 20 min, taken out, immediately cut vertically downward from the plane intersecting the line of sight, coronal sections were cut out, and sections were cut back every 2 mm, and brain sections were stained with 2% TTC. Incubate in liquid (37℃, 90 min), normal brain tissue is stained crimson, ischemic brain tissue appears white, and after flushing with physiological saline, the brain slices are quickly lined up from front to back. They were lined up, absorbed the water remaining on the surface, and photographed.

The photographs were processed by image analysis software, and the volume corresponding to the left brain and the volume of infarcts were calculated by the following formula to determine the percentage of infarcts.
Infarct volume calculation method:
V＝t (A1+A2+A3+………+An)
t is the thickness of the section and A is the infarct area.
% I = 100% x (VC-VL)/VC
%I is the percentage of infarct volume, VC is the brain volume on the control side (left hemisphere) and VL is the volume of the non-infarct region on the infarct side (right hemisphere).

2 experimental results
2.1 Effects on neurological deficit symptoms Table 1 shows the degree of neurological deficit symptoms in each group of animals. Compared with the model group, the three sublingual doses of the composition (3, 9, 27 mg/kg) and the positive control compound edaravone (3 mg/kg) all showed significant neurological deficit symptoms. Can be improved.
Table 3. Effects of sublingual combined drug of edaravone and (+)-2-borneol on neurological deficit symptoms

実験データにより、本発明にかかる舌下錠は注射剤と同等の薬効を達成できることが示された。 2.2 Effect on cerebral infarct area The effect on cerebral infarct area is shown in Table 2. All three sublingual doses of the composition (3, 9, 27 mg/kg) and the positive control compound edaravone reduce cerebral infarct area after animal ischemia-reperfusion compared to the model group be able to.
Table 2 Effect of combined drug for sublingual administration of edaravone and (+)-2-borneol on cerebral infarct area

The experimental data showed that the sublingual tablet according to the present invention can achieve a drug effect equivalent to that of an injection.

Claims (9)

A pharmaceutical composition for sublingual tablet for treating cerebrovascular disease , comprising edaravone, (+)-2-borneol and a pharmaceutically acceptable additive , wherein the pharmaceutically acceptable additive is mannitol and A pharmaceutical composition for sublingual tablets for treating cerebrovascular disease, which comprises copolyvidone . Mass ratio of 1: 5 to 5: and a 1 mannitol and copolyvidone, pharmaceutical composition according to claim 1. The pharmaceutical composition according to claim 2, wherein the mass ratio of mannitol and copolyvidone is 1:1 to 5:1. The pharmaceutical composition according to any one of claims 1 to 3, wherein the mass ratio of edaravone and (+)-2-borneol in the free base is greater than 4 and less than 10 or greater than 0.1 and less than 1. .. The pharmaceutical composition according to any one of claims 1 to 4, wherein the mass ratio of edaravone and (+)-2-borneol as a free base is 5:1. The pharmaceutical composition according to any one of claims 1 to 5, wherein the weight ratio of (+)-2-borneol to mannitol and copolyvidone is 0.1:1 to 1:1. The pharmaceutical composition according to any one of claims 1 to 5, wherein a weight ratio of (+)-2-borneol , mannitol and copolyvidone is 0.3:1 to 1:1. The drug blood concentration of edaravone reached 10 to 8000 ng/mL within about 0.1 to 10 hours after administration of sublingual tablets according to the standard unit to the patient, and the drug blood concentration of (+)-2-borneol was The pharmaceutical composition according to any one of claims 1 to 7, which reaches 1 to 200 ng/mL. The drug blood concentration of edaravone reached 50-5000 ng/mL, and the drug blood concentration of (+)-2-borneol reached 2 within about 0.1 to 6 hours after the administration of sublingual tablets per standard unit. The pharmaceutical composition according to any one of claims 1 to 7, which reaches ~150 ng/mL.

Images